In this research proposal we plan to apply biochemical and genetic approaches to the study of amino acid transport in eukaryotic cells. We are characterizing the transport systems for neutral amino acids from several mammalian cell lines. In addition, membrane vesicle preparations will be made from the cells and used for transport studies. We will continue our studies on the reconstitution of neutral amino acid transport systems from Ehrlich ascites cell plasma membranes. Cholate and other detergents are being used to solubilize the transport components from the plasma membrane. Purification of the components involved in transport is the eventual goal of the reconstitution studies. Another aim of the proposal is the study of amino acid regulation in normal and virus-transformed cells. The involvement of aminoacyl synthetases in transport regulation is being studied in Chinese hamster cell lines. Those cell lines with temperature-sensitive aminoacyl synthetases are being utilized for these studies. We also hope to determine if membrane transport changes occur during malignant transformation and, if so, whether they are early or late events in the transformation process. Finally, we plan to select transport mutants in Chinese hamster ovary cells and genetically map potential mutants to human chromosomes by cell fusion techniques forming human-hamster cell hybrids.